Studies On The Mechanism Of Massive Cell Vacuolization And Autophagy Induced By Bestrophin1 And Its Mutants

Bestrophin has long been seen as the candidate of the CACC(calcium activated chloride channel).But this was changed since the finding of Anoctamin(TMEM16A gene product)as the real CACC in year 2008.So the roles of Bestrophins in vitro become more confusing at that time.Bestrophin was also revealed to be the putative candidates of volume-regulated ion channels.Recently the crystal structure of Bestrophin was generated which revealed a Ca2+bowl.However,the successful generation of the crystal of Bestrophin homolog in bacteria revealed it was Na+channel.The roles of Bestrophin family potentially display diversity or versatility.In this report,we described the initiation of cell vacuolization and strong autophagy by a mutant of human bestrophin 1.We found vacuoles from hBestl(Y227N)were highly related to microautophagy and endoplasmic reticulum(ER),and strong macroautophagy was also seen.However,blocking macroautophagy by chemical inhibitors or by shRNAi of ATG,didn’t block vacuolization.However,enhancing macroautophagy by overexpression of LC3a reduced hBestl(Y227N)vacuole formation.hBestl(Y227N)vacuoles led to malignant consequences that made the autophagosome swelling and less acidic.Unlike the typical microautophagy of yeast,microautophagy from hBestl(Y227N)was mediated by rab7.Ion imbalance within intralysosome vesicles formed by lysosome engulfment may be one factor for the swelling of microautophagosome,but severe autophagy,irregular shape of ER network and ER calcium releasing disorder may be key factors for cell vacuolization and autophagy induced by hBestl(Y227N).